Despite the fact that fatty acids are known to be potent differentiation and pro-apoptotic agents, their mechanism of action remains poorly defined. Our laboratory has demonstrated that in vitro exposure to the fatty acids butyrate, phenylbutyrate (PB) and tributyrin (TB) results in increased cellular differentiation, decreased cell proliferation, and cell cycle arrest in colon cancer cells. Importantly, we have also observed that these fatty acids are potent inducers of apoptosis and this programmed cell death is associated with an early and dramatic decrease in DNA binding and transcriptional activity of NF-kappaB, a critical regulator of apoptosis. NF-kappaB activity is regulated by associating with one of a number of inhibitory proteins; the most widely studied being IkappaB. While the IkappaB pathway represents the most extensively studied mechanism of NF-kappaB regulation, other regulatory pathways exist. One such alternative pathway may be mediated through peroxisome proliferator-activated receptors (PPAR). PPAR are novel ligand-dependent nuclear receptors that bind fatty acids and once activated, act as transcription factors that regulate genes involved in fatty acid metabolism and cellular growth. Therefore, the central hypothesis of this proposal is that fatty acid-induced apoptosis is mediated through the inhibition of NF-kappaB in colon cancer cells. To test this hypothesis, the following experiments are proposed. The first specific aim will determine whether alterations in the constitutive NF-kappaB activity affects the sensitivity to fatty acid-induced apoptosis. The second specific aim will test the hypothesis that fatty acids inhibit NF-kappaB through regulation of the IKK/IkappaB pathway. Finally, the third specific aim will test the hypothesis that fatty acid activation of PPARy induces inhibition of NF-kappaB. Given the central role of NF-kappaB in regulating cell proliferation and apoptosis, we believe that defining the mechanism(s) of fatty acid-induced inhibition of NF-kappaB will provide important mechanistic insight into these novel and potentially clinically applicable agents.